The proposed research study will facilitate the improvement of the education and career goals of the principal investigator (PI). Moreover, the candidate will investigate the mechanisms that underlie impaired insulin secretion in type 2 diabetes mellitus (T2DM). The candidate has set up an important educative plan for the first two years (K99 phase) of the proposed study. This plan includes also courses. In addition, the candidate will be trained in scientific and professional research skills by his mentors and other experts. The candidate will perform experiments to investigate the molecular mechanisms underlying the functional role of intracellular calcium leak in the pathophysiology of T2DM and will suggest innovative, targeted therapeutic approaches for the treatment of such a disease. Upon conclusion of the mentored phase the candidate will have established a pool of data, knowledge and scientific skills that will enhance his credentials for successful transition to independence. During the K99 phase the candidate has planned to attend courses on biostatistics, grant writing/research funding, human subject protection, and responsible conduct of research. In addition, he has set up a specific training plan with his mentors and his advisors/collaborators that will provide him with scientific knowledge on ER stress/mitochondrial fitness, ?-cell dynamics, ion channels. These aspects represent useful tools that he will be able to transfer to his own laboratory. Besides the new techniques that he will acquire, the proposed project requires knowledge in areas that he has only recently been associated with, such as mitochondrial pathophysiology and metabolic signaling. Therefore, he has included in his advisory board a number of scientists with outstanding careers in these particular fields that will provide guidance of the highest possible level. All his advisors and collaborators are top-notch academic scientists (see a complete list in Personnel, in the `Budget justification' document). The candidate will also present his work at several international meetings, in order to receive input by a broad scientific audience, thereby expanding his scientific network. This will promote his transition to an independent research position and provide him a robust scientific foundation from which to apply for independent R01-level funding. The application core questions are: (1) Which is the functional role of intracellular Ca2+ leak via RyR2 in pancreatic ?-cells? (2) What i the mechanism underlying impaired glucose-stimulated insulin secretion in ?-cells harboring leaky RyR2 channels? (3) How can the leak be prevented in human islets from T2DM donors or animal models of diabetes? (4) How the prevention of intracellular Ca2+ leak can impact the pathophysiology of T2DM? To address these questions the candidate has designed an experimental plan with two main branches. The first falls into the K99 phase and is based on preliminary data of the current submission. This set of experiments aims to identify the mechanism that makes the Ca2+ leak via RyR2 channels in ?-cells a heretofore undisclosed key player in glucose-induced insulin release. The applicant will characterize both in vivo and ex vivo the functional aspects of leaky RyR2 using knock-in mice harboring genetic mutations in RyR2 that renders the channel leaky. The same mutations characterize a rare syndrome, known as catecholaminergic polymorphic ventricular tachycardia (CPVT); intriguingly, studies provided as material supporting the present application have demonstrated that most of these patients display glucose intolerance. The candidate will verify the effects of the induction of the leak in vitro on insulin release. Moreover, he will test an orally available small molecule (Rycal) in the CPVT mouse models, to see whether such pharmacological intervention, able to prevent intracellular Ca2+ leak, can affect ?-cell dynamics. The second branch of the plan falls into the R00 phase, where the candidate will determine whether the inhibition of intracellular Ca2+ leak via RyR2 improves glucose homeostasis in established models of T2DM. These models will be characterized with high throughput analysis methods and new targets may come up. Equally important, the elucidation of the mechanisms that will be investigated in the K99 phase (ER stress, mitochondria, ion channels) might also identify new targets and may suggest novel approaches for the treatment of T2DM. Ultimately, application of these strategies and interventions, as well as targeting of new factors that the analysis focused on ER Stress and Ca2+ fluxes is most likely going to indicate new directions to treat conditions of impaired glucose homeostasis. Moreover, with the scientific and professional skills acquired by the applicant during the K99 phase, the transition to independent position will be certainly facilitate. Overall, the current application will equip the candidate with novel knowledge and useful tools to continue for independent career in molecular metabolism and diabetes research. His training plan has been meticulously designed to facilitate flawless production of data in the prominent environment of Columbia University, as well as the PI's acquaintance with modern tools of biomedical research that he will transfer to his independent laboratory. Approval of his application for the K99/R00 award will result in the precise definition of novel mechanisms affecting ?-cell pathophysiology, challenging current paradigms and will thereby provide potential new therapeutics for the treatment of diabetes mellitus.